During FY16 we accomplished the following: 1. We completed electron microscopy studies of mitochrondrial content of CD4+ nave and memory cells as a function of age. We found the total mitochondrial content, as assessed by mitochondrial area, was not significantly different between cells obtained from young or old donors. Mitochrondrial levels were higher in memory CD4+ cells compared to nave CD4+ T cells. 2. To assess mitochondrial function we used the Oroboros Oxygraph. We found that CD4+ T cells from older individuals had reduced respirational capacity. Increased levels of electron transport chain proteins and reduced functionality led us to propose that defective mitochrondria were not being cleared by mitophagy. To address this hypothesis we initiated studies to examine LC3 expression by confocal microscopy. We imaged cells from 4 young and 4 old donors, and image quantitation is underway. 3. We carried out RNA-Seq analyses of PBMC cell subsets obtained from the first 25 GESTALT donors. This first lot involved 270 samples, and data evaluation is being carried out in collaboration with Dr. Zia Khan (UMD) and Alexis Battle (Johns Hopkins). 4. Proteomic studies were also carried out from the same group of donors. Data analyses are underway. 5. We collected purified PBMC subsets from the second group of 25 donors. RNA, protein and DNA methylation data collection is ongoing. 6. We continued our studies of cloning antigen-specific antibody genes from plasmablasts of rheumatoid arthritis patients. We have now accumulated a panel of 167 human monoclonal antibodies that are being tested for auto-reactive specificities in collaboration with Dr. Ben Larman (Johns Hopkins School of Medicine).